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Engineering a stripped-down bacterial drug factory

Researchers have deleted large chunks of a bacteria's genome in order to focus …

Many of the drugs we use are natural compounds or their derivatives, obtained from plants, fungi, or bacteria. Unfortunately, these organisms produce them for their own needs, and don't always make enough for us to obtain them in sufficient bulk or purity. One of the things that motivates the field of synthetic biology is the hope that we can design an organism from scratch so that it will make useful compounds like these drugs. But a paper that will be released by PNAS later this week suggests that there may be an easier way to go about things: take an existing bacteria and delete anything it doesn't need to make the drugs.

One of the challenges of engineering bacteria to produce natural compounds is that the chemicals involved in the production process—the biosynthetic pathway, as it's called—may come from many different parts of a cell's metabolism. So, for example, the biosynthetic pathway may stitch together a piece of a sugar, part of a broken-down protein, and some lipid in order to make a useful drug compound. So, it's not simply enough to identify the enzymes that catalyze the steps in a biosynthetic pathway; you have to identify the raw materials, too, and ensure that your engineered bacteria makes all of those.

There are two potential approaches to this, each with its share of drawbacks. One is the ground-up approach, most closely associated with Craig Venter, who has partnered with Exxon-Mobil to develop biofuels based on his technology. Venter's group started with a bacteria that has the smallest known genome, called Mycoplasma genitalium (yes, as the name implies, it's a genital parasite), and then deleted every gene in it to determine which were essential for it to survive in culture. He's also developed the ability to construct an artificial genome, which should allow him to replace the one that the bacteria already has.

This synthetic approach has two significant advantages: complete control, and the fact that the bacteria is only using the bare minimum of energy to stay alive, meaning any excess will be funneled into making your molecule of choice. The downside, however, is that you have to have complete knowledge of every step of the biosynthetic pathway, including every place where a chemical is siphoned off from the basic metabolism. For many natural compounds, we're not there yet.

The new paper takes an opposite approach: start with a useful bacteria, leave its entire metabolism intact, and then strip out anything you're not interested in. In this case, the authors started with a species of Streptomyces, a soil bacteria that's already in use for industrial production. The soil environment is a very competitive one, and different Streptomyces species produce a variety of useful compounds; the authors list chemicals with "antibacterial, antifungal, antiviral, and antitumor activities but also antihypertensive and immunosuppressant properties." These chemicals, which are optional for the bacteria's survival in the lab, are derived from the core metabolism, and called secondary metabolites.

Conveniently, the genes for most secondary metabolites are primarily clustered in two locations in the genome. So the authors simply deleted them, taking 1.4 Megabases (about 20 percent of the genome) out with a single targeted mutation. A few other specific biosynthetic pathways were also lopped off, leaving a Streptomyces strain with little more than its core metabolism.

They then plugged a few biosynthetic pathways back individually. For example, they were able to take a cluster of genes that produced the antibiotic streptomycin and drop it into their strain. It produced the antibiotic at higher levels than it would normally, presumably because it didn't have additional pathways to divert chemicals to. Another experiment converted the lab strain to one that produced a compound with antitumor properties. They could even control the expression of these biosynthetic pathway by manipulating the genes that normally regulate its activity.

In addition to compounds normally produced by bacteria, they were able to tweak a plant gene and get the Streptomyces to produce a compound called amorpha-4,11-diene. That's a precursor to the plant product artemisinin, which is a potent antimalarial drug.

Because so much of the basic metabolism of the bacteria is intact, the researchers were able to get away with a minimum amount of genetic engineering, and didn't need to identify every step in a biosynthetic pathway to get things to work. But, because of the deletions, the bacteria were focused on generating a single product, which allowed them to produce it more efficiently (though probably not as efficiently as one with a synthetic genome could).

In the end, both of these approaches will have their uses, and it's possible the two might ultimately be integrated into a single process, with bacteria producing an intermediate, which synthetic organisms go on to shape into the final product. However things work out, it's good to have options.

16 Reader Comments

"The downside, however, is that you have to have complete knowledge of every step of the biosynthetic pathway, including every place where a chemical is siphoned off from the basic metabolism. For many natural compounds, we're not there yet."

It's more like most or pretty much all natural compounds, we're not there yet.

It seems to me that they might also be able to use this method to create super effective vaccines, just delete all the harmful effects of the bacteria. Instead of using tiny bacterium corpses, I mean. Which seems so morbid.

If nothing else, the 'cool factor' of synthetic biology is off the charts. It's amazing to me that we're getting close to being able build up from scratch, rather than just introducing a gene or two at a time and praying that everything works mostly like we'd expect.

I'm doing computer science these days, though I've had a fair bit of exposure to biologists' toolkits due to a stint in a Pharmacology lab. I've noticed that many of my peers in CS place evolved systems on something of a pedestal, but my background has led me to have a very different view.

My view tends towards thinking that evolution has created some truly incredible hacks, but that it's one shitty engineer and is the last one you should be looking to for design inspiration. Simply put, there is no high level design going on, and trying to reverse engineer the soup in our cells is maddeningly difficult and slow going. Partly due to imprecise hardware (such as receptors are never 100% selective for "their" signalling molecule, and DNA that happily carries along meaningless mutations so long as the organism doesn't break too badly) and partly due to design by historical accident (famously, take the Panda's thumb) all biological systems look to me to be a cluster of messy hacks. Some of the hacks are incredible, but just as often, it seems incredible that it works at all.

The thought that we can possibly build from the ground up, whilst having a solid understanding of each minimal component, really gives some hope that we'll eventually have the sort of rapid progress in engineered organisms that we've had in engineered everything else. Evolution simply doesn't care how complex something is -- it's not a barrier to further manipulations. For us, any way that we could possibly reduce the complexity by a few orders of magnitude would be a godsend.

Imagine that a certain amount of "pressure" is needed to get organisms into "evolutionary potential wells" (heh heh, quantum biology?). Nature does this quite randomly at the moment infinite monkeys writing Shakespeare's plays in action. Or for a more appropriate analogy, pollen floating on the wind.

Humans, however, can help. We can grab a flower, carry it over to another flower, shake it, and cross our fingers. This has actually worked out quite well for us. But it's still too...random. We're pretty smart now, with all this genome stuff.

Why carry the flower over, if you can tell what genes needs to be added, modified, or removed? If it creates the same product as 10,000 attempts at cross-breeding, is it any less natural than shaking flowers on each other?

It makes sense to start with an organism that is close to a desired set of traits, and then use deletion and addition to push it over the potential hump, guiding it carefully towards our desired well without the need for random mutations and recombinations to accumulate over generations. The new organism might be stable and competitive, probably much more so than bottom-up. Unfortunately, evolution might have bigger plans for our new organism, so we may have to periodically re-populate with pre-volved organisms to keep us from sliding around the well too much.

Perhaps...too competitive? Now that I think about it, inability to survive in the wild could be an attractive selling point of the bottom-up approach...

"In the end, both of these approaches will have their uses, and it's possible the two might ultimately be integrated into a single process"

Like everything else the timing and exactly what will work out is uncertain. Craig Vetner seems to have run into some unexpected problems with his approach. But it seems very likely that these steps are just the beginning of a trend that over the next fifty to one hundred years will have a larger impact on human life than the industrial revolution or the development of the computer.

Originally posted by dreemernj:"Hey guys, I have a great idea! I'm going to make a super powerful version of a genital parasite. What could possibly go wrong?"

Not sure of the sarcasm factor on this one, but these are not more 'powerful' strains at all. They're specialized for drug production by (in the case of Venter's M. genitalium system) stripping out everything non-essential in the genome, so they are unlikely to be hardy strains outside of the gentle environment of lab cultures.

It's also interesting to see bacteria (or CHO cells or whatever) used as drug factories in general--not just for stuff they craft naturally because they compete in soil, but for introduced genes/plasmids to make even human protein. The thought that say, Micera, is grown in big vats of hamster ovary cells and then slurped out of there is pretty amazing.

Tack on some surface receptors so it can mimic a person's cells without being attacked by their immune system, and some way to sustain itself while in the human body, then you can just inject the little buggers into someone.

For a straight vaccine, that sounds impractical. But when you think about blood oxygen scrubbers allowing a person to stay under water longer, or reactive structures that would sit dormant until a certain toxin was introduced into the body, then it makes sense.

In programming, I think we take it for granted we can just toss instructions around on the computer right and left so easily, we think we're that far ahead in bio-engineering, too. But bio-engineering is really damn complicated. That's why we're still cutting folks open to do surgery (which is quite barbaric given that it's the 21st century) instead of tossing them into a bacta-tank ala Star Wars.

Wow, I almost feel bad for the bacteria. I can just imagine tens of thousands of the poor things strapped to metal tables in a huge, sterile room with tubes and pipes running through them with a mad scientist looking down at them expressionlessly, heedless of the agony their twisted, mutated forms are feeling. Good thing they're too small to be cute and cuddly.

Originally posted by kumquat:Wow, I almost feel bad for the bacteria. I can just imagine tens of thousands of the poor things strapped to metal tables in a huge, sterile room with tubes and pipes running through them with a mad scientist looking down at them expressionlessly, heedless of the agony their twisted, mutated forms are feeling. Good thing they're too small to be cute and cuddly.

Considering the range of unpleasant things that various bacteria have been doing to humans for millennia, I think that a little turn-about is not only fair play, but long overdue. ;-)